Enamel fluorosis is a defect in enamel development seen after exposure to excess fluoride during tooth formation. Much remains to be learned about the mechanisms by which fluoride affects enamel formation, and in particular the effect of fluoride on ameloblast function. Studies of human populations and mouse strains suggest that a genetic component to individual susceptibility to enamel fluorosis. In these studies we will use mouse models known to be either fluoride sensitive (A/J) or fluoride resistant (129P3/J) to identify genes responsible for this variable response to fluoride. The overall hypothesis that will be tested in these studies is that enamel fluorosis results from factors that alter the relative levels of amelogenin and MMP-20 during enamel development. This hypothesis will be tested by the following Specific Aims. Specific Aim 1: To determine the correlation between the expression of the candidate genes and severity of fluorosed enamel in the A/J and 129P3/J mouse strains. Specific Aim 2: To determine whether fluoride differentially effects phosphorylation of JNK and cJun, in A/J and 129P3/J mouse strains. Specific Aim 3: To compare the regulatory regions of the amelogenin gene in both A/J and 129P3/J mouse strains and determine how the differences relate to susceptibility/resistance to Dental fluorosis. We will use laser microdissection microscopy to separate secretory and maturation stage ameloblasts from incisors of mice given fluoride in drinking water. Real-time PCR will be used to quantitate relative expression of amelogenin, MMP-20, and other candidate genes to identify variable response to fluoride in the genetically diverse mouse strains. Genetic differences in regulators of MMP-20 will be further investigated by measuring relative differences in JNK/c-Jun synthesis and phosphorylation in the two mouse models. Polymorphisms in the amelogenin gene will be investigated by comparing promoter sequences in the two mouse models. We anticipate that these studies will result in the identification of candidate genes that will form the basis an R01 application to further determine genetic susceptibility to fluorosis in humans.